3-amino-flavanones

ABSTRACT

IN WHICH R1 IS HYDROGEN, R2 IS OH OR O (LOWER)ACYL, OR R1R2 TAKEN TOGETHER ARE O, R3 IS H OR (LOWER)ALKYL, R4 IS H OR (LOWER)ALKYL OR NR3R4 IS A HETEROCYCLIC MOIETY OF SIX MEMBERS, X IS A HALOGEN, (LOWER)ALKYL OR (LOWER) ALKOXY, AND Y IS A HALOGEN, (LOWER)ALKYL, OR (LOWER)ALKOXY, A PROCESS FOR PREPARING SUCH COMPOUNDS, AND A PROCESS FOR INHIBITING THE GROWTH OF MICROORGANISMS.   2-(Y-C6H4-),3-(R3-N(-R4)-CH2-),4-R1,4-R2,X-CHROMAN   A GROUP OF FLAVANONE DERIVATIVES OF THE STRUCTURAL FORMULA

United States Patent IC@ 3,753,985

Patented Aug. 21, 1973 of six members, X is a halogen, (lower)alkyl or(lower) 3,753,985 alkoxy, and Y is a halogen, (lower)alkyl or (lower)alk- 3-AM1N O-FLAVAN ONES oxy. Preferably, in this invention the(lower)acyl, (lower) John Joseph Gavin, Hamden, and Hermaii HenryWalchlr,

Jr. West Haven Conn. and Dale drian Stautfer t z Ind. assignors MilesLaboratories, Inc: 5 a oms inclusive. The heterocyclrc moieties areadvanta El" Ind geously selected from morpholino, piperidino,piper'azino No a g Filed Dec. 28 1970,, Ser- No. 102 181 and similar SIXmembered rings. The halogen included In CL C077! 7 /20 in this structureis advantageously selected from chlorine, U.s. Cl. 260-247.7 G 3 Claimsbromine or fluorine- J I 10 The compounds of this invention may beprepared according to a series of chemical 'reactions'inwhich finalABSTRACT OF THE DISCLOSURE products may be selectively separated fromthe reaction A group of fiavanone derivatives of the structuralformixture or further reacted without separation to form mula otherfinal products. This sequence is initiated by a Man- Y nich reactioninvolving a fiavanone, formaldehyde and a primary, secondary or tertiaryamine. The resulting 3-sub- H stituted flavanone may then be separatedor the reaction X F continued by a reduction of this product in thepresence of Re a suitable catalyst to form a 3-substituted flavan-4-ol.This CH final product may be separated if desired or be allowed to R R22 remain in solution and be acylated to form a 3-aminoalkyl-4-acyloxyfiavan. Any of the above final products may be in which R1 ishydrogen R2 is OH or O (lower)acyl or reacted according to knownprocedures to form acid ad- Rl z taken to ether are 0 R3 is H or loweralk 1 R4 dition salts thereof and preferably to form pharmacologis H or(lowerialkyl or NR R is a heterc cyclic moi e ty of y acceptable aclfladdmon Salts thereof This y six members, X is a halogen, (lower)alkyl or(lower) them. may be .graphlcauy represented by the followmg alkoxy, andY is a halogen, (lower) alkyl, or (lower)alkchemlcal equanoni oxy; aprocess for preparing such compounds; and a proc- Y ess for inhibitingthe growth of microorganisms. Q

@fiK/ X BACKGROUND OF THE INVENTION ECHO It has been known that variouschemicals may be used to inhibit the growth of microorganisms such asyeast and molds. Although it would be desirable to have a single 0chemical that would inhibit the growth of all undesirable Ymicroorganisms, such a chemical has not been available. Therefore, toeffectively inhibit and combat infestations o of these microorganisms,it has been the practice to com- H reduction bine several chemicals toprovide the desired relief. X Such mixtures of chemicals are oftensuccessful in their H R3 attack upon these microorganisms. However, sidereac- Y CHZN/ tions may develop from sensitivity to one or more of thechemicals included in the mixture. Also, strains of yeasts and molds maydevelop that are resistant to the chemicals Y which are repeatedly usedin the treatment thereof, thereby decreasing or eliminating theinhibition capabilities of 0 these chemicals. X H E (I?) SUMMARY OF THEINVENTION W z +1) 0 im-H mo 2g This invention relates to a novel seriesof chemical compounds and a new process for inhibiting the growth of HOmicroorganisms. Also, this invention relates to a process for thepreparation of the novel compounds of this inven- Y tion. Moreparticularly, this invention relates to a series Q of flavanonederivatives which has been found especially 0 useful for inhibiting thegrowth of yeasts and molds such H as Trichophyton mentagrophytes andCandida albicans. X

Compounds of this invention may be represented by H the generalstructural formula: CHZN X In this equation Z and W are halogens whichmay be the H Ra same or difierent and m is an integer from 1 to 4inclusive. 1 R4 The growth of microorganisms is effectively inhibitedaccording to the process of this invention by exposure of in which R ishydrogen, R is OH or 0(lower) acyl, or the microorganism to a compoundselected from the group R R taken together are 0, R is H or(lower)alkyl, R 7 described above. The means by which the organism isinis H or (lower) alkyl or NR R is a heterocyclic moiety hibited is notconsidered critical and will depend upon the alkyl and (lower)alkoxyconsist of from 1 to 5 carbon a relstthslrsa i t e.a eaQtinfestatio and.. .t.lie.-- hpip num. xide c a y an 200 of methanol.

host which is affected thereby, Typically, inhibition m be achieved bytopical application of compounds of this invention. In other situationsa systemic action may be achieved with an'oralor injectable dosagefform.Typical- BQDIMEN'ITSSYN t amma es; e ne A mixture of iiavanone (112.1g., 0.5 mole), morpholine hydrochloride (61.6 g., 0.5 mole), 95% para-'formaldehyde (31.6 g., 1 mole), 250ml. ofZ-propanol and 25 drops ofconcentrated hydrochloric acid was heated to reflux. After about minutesmost of the solid material had dissolved. Then crystals began to form,and after 5-10 minutes a copious precipitate was present. The mixturewas cooled and diluted with dry ether. The salt was collected, washedwith ether and dried. The crude material was agitated for 10 minuteswith 500 ml. of boiling 2-propanol. Much of the solid material remainedundissolved. The white salt was filtered from the hot mixture and washedwith 2-propanol. Yield (88.3 g.), M.P. 172-3 C.

The filtrate and washings were concentrated by evaporation, and theresidue was diluted with dry ether. The solid material thus obtained wasmixed with 200 ml. of hot Z-propanol. The insoluble solid material wascollected and washed with 2-propanol. This second crop of the whitecrystalline salt amounted to 17.2 g. M.P. 171- 2 C. Total yield 105.5 g.

Analysis.Calcd. for C H NO -HCl (percent): HCl, 10.13, N (basic), 3.89;N (total), 3.89. Found (percent): HCl, 10.05; N (basic), 3.89; N(total), 3.89.

EXAMPLE 2 3- (dimethylaminomethyl) -fiavanone hydrochloride A mixture offiavanone (44.9 g., 0.2 mole), dimethylamine hydrochloride (16.3 g., 0.2mole), 95% paraformaldehyde (9.5 g., 0.3 mole), 100 ml. of Z-propanolwas shaken under an initial hydrogen pressure of 50 pounds. The mixturewas shaken overnight and somewhat more than the theoretical quantity ofhydrogen required to reduce the carbonyl group was absorbed. Thecatalyst was gremoved by filtration and washed with Z-propanol.Thefilhesolvents were removed from the filtrate and washings bydistillation'under reduced pressure. The

3 synupy residueawas'taken up in 200 ml. of water, and the solution waswashed by shaking with a little ether. The fre' -bas'e was liberated bythe addition of excess potassium carbonate. The solid material wascollected, washed v, with water and recrystallized from aqueous ethanol.Yield j sis.Calcd. for C H NO (percent): N (basic), N (total), 4.94.Found (percent): N (basic), 4.91; N (total),'t i; 96

EXAMPLE 4 1 3T-(dimethylaminomethyl)flavan-4-olhydrochloride The freebasic (12 .5 g.) prepared according to Example 3 was dissolved in 100ml. of warm 2-propanol, and

-- 'an ex'ce'ssof ethereal hydrogen chloride was added. Then and 10drops of concentrated hydrochloric acid was,

heated under reflux for 1 hour. Then another 4.8 g. (0.15 mole) ofparaformaldehyde was rinsed in the 25 ml. of 2-propanol, and the heatingwas continued for 2 hours longer. The solvent was removed bydistillation under reduced pressure and the syrupy residue was shakenwith 150 ml. of ether and 150 ml. of water. The aqueous portion wasseparated and treated with excess potassium carbonate. The syrupyjree:base-was taken-up .in-"ether, .1: and the extract was dried. overmagnesium sulfate; The.

dried ether solution was treated with an excess of ethereal:

hydrogen chloride. A sticky salt separatedpout which was-5" EXAMPLE '3'"3-(dimethylaminomethyl)flavan-4-ol A mixture ofS-(dImethylaminonithyl)flavanone 'hythe mixture was diluted with dryether. The salt was collected, Washed with ether and recrystallized froma mixture of methanol and 2-propanol. The white crystals amounted to12.3 g. The filtrate and washings were concentratedby evaporation anddiluted with dry ether to give -an additional 1.2 g. of the salt. Yield13.5 g., M.P.

Analysis.Calcd. for C H NO -Hcl (percent): HCl, 11.40; N (basic), 4.38;N (total), 4.38. Found (percent):

'HCl, 11.30; N (basic), 4.27; N (total), 4.42.

EXAMPLE 5 3-(4-morpholinylmethyl)flavan-4-ol A mixture of3-(4-morpholinylmethyl)flavanone hydrochloride (16.0 g., 0.045 mole)(cf. Example 1), 0.2 g. of platinum oxide catalyst and 200 ml. ofmethanol was shaken under an initial hydrogen pressure of 50 pounds.After shaking for 24 hours, slightly more than theoretical quantity ofhydrogen required to reduce the carbonyl group was taken-up. Thecatalyst was removed by filtration and washed with 2-propanol. Thesolvent was removed from the filtrate and washings by distillation underreduced pressure. The syrupy residue was taken-up in 200 ml. of water,and the solution was washed by shaking with a little ether. Then excesspotassium carbonate was added. The solid free base was collected, washedwith water and recrystallized from aqueous ethanol. Yield (11.0 g.),M.P. 182-3 C.

.. Analysis.-Calcd. for C H NO (percent): N (basic), 4.31; N (total),4.31. Found (percent): N (basic), 4.25; N (total), 4.33.

. EXAMPLE 6 '3- 4-morpholinylmethyl) flavan-4-ol hydrochloride :EXAMPLE73-(4-morpholinylmethyl)-4-propionoxyfiavan hydrochloride 3-.(4rnorpholinylmethyl)flavan-4-ol (14.9 g., 0.046 mole) (cf. Example 5) wasdissolved in 100 m1. of chloroform and propionyl chloride (8.5 g., 0.092mole) was drochloridfi 2;. 0-043 mole) EXamPle added. The solutionbecame quite warm. After the mixture PHARMACOLOGICAL EVALUATION EXAMPLE8 The ability of the comounds of this invention to inhibit the growth ofTrichophyton mentagrophytes was determined in the following manner: Aspore suspension was prepared and standardized to contain approximately1,000,000 spores per milliliter in 0.9% saline. To each of a series oftubes containing 0.9 ml. of serial two-fold dilutions of the compound tobe tested in Sabourauds broth, 0.1 ml. of the above suspension wasadded. The Sabourauds broth had the following composition, ex pressed ingrams per liter of distilled water.

TABLE 1 Pancreatic digest of casein 5.0 Peptic digest of animal tissue5.0 Dextrose 20.0

Having a final pH of 5.7 $0.1.

The dilution series ranged downward from 50 mcg. per ml. The tubes wereincubated at 37 C. and observed for the presence of or absence of growthat the time intervals set forth in Table 2. The endpoint was taken asthat tube in which growth was completely inhibited as evidenced byabsence of turbidity. Control tubes containing standard antibiotics andtubes without inhibitory substances were included for comparativepurposes.

TABLE 2 Minimum inhibitory concentration, meg/ml.

Time, hours; ';.'..'.;.".'.;'...;;1-;. 24 48 72 Compound:

3-(tmorpholinylmethyl)-flavanone hydrochloride (Compound A) .5 25 50 503- (dimehtylaminomethyl) -flavanone hydrochloride (Compound B) 303-(dimethylaminomethyl)-fiavan-4-ol hydrochloride (Compound 0) 50 Theminimum inhibitory concentration of Griseofulvin for this sporeconcentration at 72 hours was 1.9-2.1 meg/ml.

The eifect observed in this example demonstrated that the compounds ofthis invention are effective inhibitors of the growth of microorganisms.This is supported by the comparison of the minimum inhibitoryconcentration of these compounds with the widely used medicationGn'seofulvin.

EXAMPLE 9 The ability of the compounds of this invention to inhibit thegrowth of Candida albicans was determined in the following manner. Acell suspension was prepared and standardized to contain approximately1,000,000 cells per milliliter in 0.9% saline. 0.1 ml. of thissuspension was added to each of a series of tubes containing 0.9 ml. ofserial two-fold dilutions of the compound being tested in Eugon Brothhaving the following composition expressed in grams per liter ofdistilled water.

6 TABLE 3 Pancreatic digest of casein 15.0 Papaic digest soya meal 5.0Sodium chloride 4.0 Sodium sulfite 0.2 L-cystine 0.7 Dextrose 5.5

Having a final ph of 7.0:01.

The dilution series ranged downward from 50 mcg. per ml. The tubes wereincubated at 37 C. and observed for the presence or absence of growth atthe time intervals set forth in Table 4. The endpoint was taken as thattube in which growth was completely inhibited as evidenced by theabsence of turbidity. Control tubes containing standard antibiotics andtubes without inhibitory substances were included for comparison.

TABLE 4 Minimum inhibitory concentration, meg/ml Time, hours 24 48 72 96Compound:

The minimum inhibitory concentration of Nystatin for this cellconcentration at 72 hours was 2-10 meg/ml.

The eflfects observed in this example demonstrated that the compounds ofthis invention are eifective inhibitors of the growth of microorgaisms.This is particularly true when the minimum inhibitory concentration ofthe new compounds is compared with widely used medication Nystatin.

EXAMPLE 10 The oral toxicity of the compounds of this invention wasdetermined by administration thereof to rats. Groups of rats wererandomly selected and respectively received, orally, graduated doses ofthe new compounds in a saline solution. Following administration, theanimals were observed for side eifects and death. From the observeddeaths, an LD (dose at which death occurred in 50% of the animalstested) was calculated which is set forth in Table 5. A control group ofrats received a corresponding quantity of saline solution.

EXAMPLE 11 The toxicity of these compounds when administratedintraperitoneally was determined in mice. Groups of mice were randomlyselected and respectively received, intraperitoneally, graduated dosesof the new compounds in a saline solution. An equal quantity of salinesolution was administered to a group of mice for control purposes. Afteradministration, the animals were observed for side reactions and death.From the observed deaths an m was calculated which is set forth in Table5.

TABLE 5 Eat (oral) Mouse (i.p.) Compound mgJkg. mgJkg:

